Monolithic capillary column technologies

Capillary Column Technology Continuous Polymer Monoliths... 

The main bottleneck in the further development of CEC is related with the state of the art of the column manufacturing processes and the robustness of the columns/instrumentation. Moreover, evidence to demonstrate reproducibility of separations from column to column still has to be established. The formation of bubbles in the capillaries due to the Joule heating and variations in EOF velocity on passing from the stationary phase through the frit and into the open tube is still very challenging in packed column CEC. A way to overcome this problem is to use monolithic columns or apply open tubular CEC [108]. Currently, many efforts are placed in improving column technology and in the development of chip-CEC [115] as an attractive option for lab-on-a-chip separations. 

Several research groups used another interesting column technology as an alternative to the modification of the capillary surface. This method is inherited from the field of electrophoresis of nucleic acids and involves capillaries filled with solutions of linear polymers. In contrast to the monolithic columns that will be discussed later in this review, the preparation of these pseudostationary phases need not be performed within the confines of the capillary. These materials, typically specifically designed copolymers [85-88] and modified den-drimers [89], exist as physically entangled polymer chains that effectively resemble highly swollen, chemically crosslinked gels. 

This technology was extended to the preparation of chiral capillary columns [ 138 -141 ]. For example, enantioselective columns were prepared using a simple copolymerization of mixtures of O-[2-(methacryloyloxy)ethylcarbamoyl]-10,11-dihydro quinidine, ethylene dimethacrylate, and 2-hydroxyethyl methacrylate in the presence of mixture of cyclohexanol and 1-dodecanol as porogenic solvents. The porous properties of the monolithic columns can easily be controlled through changes in the composition of this binary solvent. Very high column efficiencies of 250,000 plates/m and good selectivities were achieved for the separations of numerous enantiomers [140]. 

Current research in CEC involves the use of monolith capillaries, which are fritless, packed capillaries having stationary phase bound to the capillary wall. Using porous polymer monoliths, the retention of a packed column can be found in an open tubular capillary. In general, CEC remains unsettled. Frit technology is unreliable and research into monolithic capillaries is still a work in progress. Recent progress in CEC can be found in the reviews by Colon and co-workers. 

The need of column configurations and surface chemistries especially designed for CEC is now generally appreciated and novel approaches to improve the column technology for CEC—MS applications include the use of monolithic stationary phases [109,110], open-tubular capillary columns [86] and chip technology [111]. These configurations are currently under detailed investigation and the future will have to prove their applicability in routine analysis. 

Separation columns are the heart of any CEC techniques. In order for CEC to become a versatile analytical technique, it is important to have columns with good stability, high EOF velocity generation, and affinity for different analytes. To this end, several different types of capillary columns have been developed for the separation of biomolecules. In this section, a summary of recent advances in column technology, including open tubular columns, duplex columns, and monolithic columns, will be presented. A comparison of different column configurations is shown in Figure 13. 

Monolithic technology first originated as an alternate technique to fabricate capillary columns. It is a radical departure from packed-column technology and uses in situ polymerization to form a continuous bed of porous silica24 inside the fused silica capillaries. Because end frits are problematic in packed capillaries, this new approach eliminates this problem since no end frits are required for monoliths. For years, capillary monoliths remained a scientific tool for academic research. 

Several groups used sol-gel transition to immobilize the beads packed in a capillary. For example, Dulay et al. [102] packed a slurry of ODS beads in tetraethylorthosilicate solution and heated it to 100 °C to achieve the sol-gel transition and create the monolithic structure shown in Fig. 17. This technology is extremely sensitive and even a small deviation from the optimal conditions leads to cracks in the monoliths and a rapid deterioration in the column performance. However, even the best efficiency of 80,000 plates/m achieved with these column was relatively low. Henry et al. modified the original procedure and increased the efficiencies to well over 100,000 plates/m [103,104]. 

Silica-based monolithic columns (Figure 9) are generally prepared using sol-gel technology. This involves the preparation of a sol solution and the gelation of the sol to form a network in a continuous liquid phase within the capillary. The precursors for the synthesis of these monoliths are normally metal alkoxides that react readily with water. The most widely used are alkoxysilanes such as tetramethoxysilane (TMOS) and TEOS. 

A wide variety of approaches are currently being used in the fabrication and technology of columns for capillary electrochromatography (CEC). Continuous polymer bed, or monolithic columns (see Section 3.4), manufactured by in-situ polymerization within the columns, have been used in numerous application areas and have been shown to be highly efficient. In a second approach, a sol-gel process is employed to form a silica xerogel within the capillary, followed by bonding of the stationary-phase group alternatively, the separation medium itself may be polymerized in situ. 

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